A generic sequencing based typing approach for the identification of HLA-A diversity

Hum Immunol. 1997 Oct;57(2):120-8. doi: 10.1016/s0198-8859(97)00204-8.

Abstract

Sequencing Based Typing (SBT) is a generic approach for the identification of HLA-A polymorphism. This approach includes the high resolution typing of the HLA-A broad reacting groups, HLA-A subtypes and will identify new alleles directly. The SBT approach described here uses a locus specific amplification of DNA from exon 1 to exon 5. The resulting 2,022 bp PCR product serves as a template for the subsequent sequencing reactions. Amplification is followed by direct sequencing of exons 2, 3 and 4 in both orientations with fluorescently labeled primers to define all polymorphic positions leading to a high resolution typing result. In this study the sequence of exons 2 and 3 of a panel of 49 cell lines was determined. In addition, the exon 4 region of 35 cell lines was also sequenced to evaluate the exon 4 polymorphism. The HLA-A type of most of the cells could be identified by sequencing only exons 2 and 3. However, the sequence of exon 4 was required to discriminate A*0201 from A*0209 and A*0207 from A*0215N. In this panel, an identical new "HLA-A*0103" was identified in two Caucasian samples.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alleles
  • Base Sequence
  • Cloning, Molecular
  • Exons
  • Genetic Carrier Screening
  • HLA-A Antigens / genetics*
  • HLA-A Antigens / immunology
  • Histocompatibility Testing / methods*
  • Humans
  • Molecular Sequence Data
  • Oligonucleotide Probes / genetics
  • Polymerase Chain Reaction / methods*
  • Polymorphism, Genetic
  • Sequence Alignment
  • Sequence Analysis, DNA

Substances

  • HLA-A Antigens
  • Oligonucleotide Probes

Associated data

  • GENBANK/Y14624